There have been numerous clinical trials involving various types of oligonucleotides including ‘classic’ antisense (AS), siRNAs, and splice switching oligonucleotides (SSOs), testifying to the immense interest in this broad therapeutic approach. Nonetheless, oligonucleotide-based therapies face a key problem regarding the inefficient access of oligonucleotides to their sites of action in the nucleus or cytosol of tissue cells. This problem has meant that large doses must be given to attain therapeutic effects thus risking drug-related toxicities, or that complex delivery systems such as cationic lipid or polymer nanoparticles must be used thus creating toxicity and biodistribution problems associated with the delivery system itself. Therefore it is clear that the discovery of alternative strategies to enhance the access of oligonucleotides to their intracellular targets will have substantial value for oligonucleotide-based pharmacology and therapeutics.
Oligonucleotides are usually internalized via endocytosis and then traffic through various membrane-bound vesicular compartments. Cells employ multiple distinct endocytotic uptake mechanisms including the ‘classic’ clathrin pit pathway, the caveolar pathway, one or more caveolin and clathrin-independent pathways, and macropinocytosis. Initial uptake is followed by trafficking into a variety of endomembrane compartments including early/sorting endosomes, late endosomes/multi-vesicular bodies, lysosomes, and the trans-Golgi network (TGN). Most of the oligonucleotide accumulated in cells remains sequestered in endomembrane vesicles and is pharmacologically inert, but a small fraction escapes to the cytosol and nucleus to permit activity. Recently we, and others, have found that the route of uptake and pathway of intracellular trafficking can have a strong effect on the pharmacological activity of the oligonucleotide; there are productive and less productive pathways. These observations suggest that if it were possible to influence the intracellular trafficking of oligonucleotides, and their release from endomembrane compartments, one might be able to substantially enhance their pharmacological effects and/or the physiological activity thereof. See, e.g., R. Juliano et al., Small Molecules that Enhance the Activity of Oligonucleotides, PCT Application WO 2013/123217 (22 Aug. 2013).